Staphylococcus aureus is the causative agent for a wide range of human diseases, especially suppurative infections. S aureus is also the single most common cause of hospital acquired (nosocomial) infections in American hospitals today. Nosocomial infections are of extreme concern, because of the increased admittance of immunocompromised patients in American hospitals, particularly patients with AIDS. This problem is further aggravated by the occurrence of multiple drug resistances in S aureus, with few effective antibiotic regimens available. Recent hospital isolates of S aureus were resistant to all known antibiotics and the development of new targets for antibiotic therapy is therefore urgently needed. A variety of surface proteins is responsible for staphylococcal virulence, but the mechanism by which these proteins anchor to the surface is not understood. In this proposal it is shown that protein A, another surface protein, is anchored in the staphylococcal cell wall. The signal for cell wall anchoring is encoded in three C-terminal sequence elements of protein A: a LPXTGX sequence motif, a C-terminal hydrophobic domain, and a charged tail. These three sequence elements are conserved in more that 30 different surface proteins from different gram-positive species suggesting that these proteins may anchor in a similar manner. In this proposal, the sorting signal sufficient for cell wall anchoring of protein A will be characterized with mutant and hybrid proteins. The linkage between protein A and the staphylococcal peptidoglycan will be studied by direct methods of chemical analysis. Furthermore, mutagenesis experiments will screen for the genetic determinants responsible for cell wall sorting of protein A in S aureus. Cell wall anchoring of surface proteins may be specific for gram-positive bacteria and could therefore present a new target for antibiotic therapy.